Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures
Abstract Background Esters are versatile chemicals and potential drop-in biofuels. To develop a sustainable production platform, microbial ester biosynthesis using alcohol acetyltransferases (AATs) has been studied for decades. Volatility of esters endows high-temperature fermentation with advantage...
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| Format: | Article |
| Language: | English |
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BMC
2019-10-01
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| Series: | Biotechnology for Biofuels |
| Subjects: | |
| Online Access: | http://link.springer.com/article/10.1186/s13068-019-1583-8 |
| _version_ | 1811255839592284160 |
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| author | Hyeongmin Seo Jong-Won Lee Sergio Garcia Cong T. Trinh |
| author_facet | Hyeongmin Seo Jong-Won Lee Sergio Garcia Cong T. Trinh |
| author_sort | Hyeongmin Seo |
| collection | DOAJ |
| description | Abstract Background Esters are versatile chemicals and potential drop-in biofuels. To develop a sustainable production platform, microbial ester biosynthesis using alcohol acetyltransferases (AATs) has been studied for decades. Volatility of esters endows high-temperature fermentation with advantageous downstream product separation. However, due to the limited thermostability of AATs known, the ester biosynthesis has largely relied on use of mesophilic microbes. Therefore, developing thermostable AATs is important for ester production directly from lignocellulosic biomass by the thermophilic consolidated bioprocessing (CBP) microbes, e.g., Clostridium thermocellum. Results In this study, we engineered a thermostable chloramphenicol acetyltransferase from Staphylococcus aureus (CATSa) for enhanced isobutyl acetate production at elevated temperatures. We first analyzed the broad alcohol substrate range of CATSa. Then, we targeted a highly conserved region in the binding pocket of CATSa for mutagenesis. The mutagenesis revealed that F97W significantly increased conversion of isobutanol to isobutyl acetate. Using CATSa F97W, we demonstrated direct conversion of cellulose into isobutyl acetate by an engineered C. thermocellum at elevated temperatures. Conclusions This study highlights that CAT is a potential thermostable AAT that can be harnessed to develop the thermophilic CBP microbial platform for biosynthesis of designer bioesters directly from lignocellulosic biomass. |
| first_indexed | 2024-04-12T17:30:57Z |
| format | Article |
| id | doaj.art-43a8bb5777f54737a88d54acf2d52e8d |
| institution | Directory Open Access Journal |
| issn | 1754-6834 |
| language | English |
| last_indexed | 2024-04-12T17:30:57Z |
| publishDate | 2019-10-01 |
| publisher | BMC |
| record_format | Article |
| series | Biotechnology for Biofuels |
| spelling | doaj.art-43a8bb5777f54737a88d54acf2d52e8d2022-12-22T03:23:08ZengBMCBiotechnology for Biofuels1754-68342019-10-0112111310.1186/s13068-019-1583-8Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperaturesHyeongmin Seo0Jong-Won Lee1Sergio Garcia2Cong T. Trinh3Department of Chemical and Biomolecular Engineering, The University of TennesseeBredesen Center for Interdisciplinary Research and Graduate Education, The University of TennesseeDepartment of Chemical and Biomolecular Engineering, The University of TennesseeDepartment of Chemical and Biomolecular Engineering, The University of TennesseeAbstract Background Esters are versatile chemicals and potential drop-in biofuels. To develop a sustainable production platform, microbial ester biosynthesis using alcohol acetyltransferases (AATs) has been studied for decades. Volatility of esters endows high-temperature fermentation with advantageous downstream product separation. However, due to the limited thermostability of AATs known, the ester biosynthesis has largely relied on use of mesophilic microbes. Therefore, developing thermostable AATs is important for ester production directly from lignocellulosic biomass by the thermophilic consolidated bioprocessing (CBP) microbes, e.g., Clostridium thermocellum. Results In this study, we engineered a thermostable chloramphenicol acetyltransferase from Staphylococcus aureus (CATSa) for enhanced isobutyl acetate production at elevated temperatures. We first analyzed the broad alcohol substrate range of CATSa. Then, we targeted a highly conserved region in the binding pocket of CATSa for mutagenesis. The mutagenesis revealed that F97W significantly increased conversion of isobutanol to isobutyl acetate. Using CATSa F97W, we demonstrated direct conversion of cellulose into isobutyl acetate by an engineered C. thermocellum at elevated temperatures. Conclusions This study highlights that CAT is a potential thermostable AAT that can be harnessed to develop the thermophilic CBP microbial platform for biosynthesis of designer bioesters directly from lignocellulosic biomass.http://link.springer.com/article/10.1186/s13068-019-1583-8Alcohol acetyltransferaseThermostabilityChloramphenicol acetyltransferaseIsobutyl acetateEstersConsolidated bioprocessing |
| spellingShingle | Hyeongmin Seo Jong-Won Lee Sergio Garcia Cong T. Trinh Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures Biotechnology for Biofuels Alcohol acetyltransferase Thermostability Chloramphenicol acetyltransferase Isobutyl acetate Esters Consolidated bioprocessing |
| title | Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures |
| title_full | Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures |
| title_fullStr | Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures |
| title_full_unstemmed | Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures |
| title_short | Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures |
| title_sort | single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by clostridium thermocellum at elevated temperatures |
| topic | Alcohol acetyltransferase Thermostability Chloramphenicol acetyltransferase Isobutyl acetate Esters Consolidated bioprocessing |
| url | http://link.springer.com/article/10.1186/s13068-019-1583-8 |
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